A one health approach versus Acanthamoeba castellanii, a potential host for Morganella morganii / Un enfoque de salud único versus Acanthamoeba castellanii, un huésped potencial de Morganella morganii

Acanthamoeba castellanii, known as the “Trojan horse of the microbial world,” is known to host a variety of microorganisms including viruses, yeasts, protists, and bacteria. Acanthamoeba can act as a vector and may aid in the transmission of various bacterial pathogens to potential hosts and are found in a variety of places, thus impacting the health of humans, animals, and the environment. These are interconnected in a system known as “one health.” With the global threat of antibiotic resistance, bacteria may avoid harsh conditions, antibiotics, and disinfectants by sheltering within Acanthamoeba. In this study, Acanthamoeba castellanii interaction with Morganella morganii, a Gram-negative bacterium was studied. Escherichia coli K1 interaction with Acanthamoeba was carried out as a control. Association, invasion, and survival assays were accomplished. Morganella morganii was found to associate, invade, and survive within Acanthamoeba castellanii. Additionally, Escherichia coli K1 was also found to associate, invade, and survive within the Acanthamoeba at a higher number in comparison to Morganella morganii. For the first time, we have shown that Morganella morganii interact, invade, and survive within Acanthamoeba castellanii, suggesting that Acanthamoeba may be a potential vector in the transmission of Morganella morganii to susceptible hosts. Taking a one health approach to tackle and develop disinfectants to target Acanthamoeba is warranted, as the amoebae may be hosting various microbes such as multiple drug-resistant bacteria and even viruses such as the novel coronavirus.(AU)

High-Pressure Inactivation of Histamine-Forming Bacteria Morganella morganii and Photobacterium phosphoreum.

ABSTRACT: The effects of high hydrostatic pressure (HHP) treatments on histamine-forming bacteria (HFB) Morganella morganii and Photobacterium phosphoreum in phosphate buffer and tuna meat slurry were investigated using viability counting and scanning electron microscopy. The first-order model fits the destruction kinetics of high pressure on M. morganii and P. phosphoreum during the pressure hold period. The D-values of M. morganii (200 to 600 MPa) and P. phosphoreum (100 to 400 MPa) in phosphate buffer ranged from 16.4 to 0.08 min and 26.4 to 0.19 min, respectively, whereas those in tuna meat slurry ranged from 51.0 to 0.09 min and 71.6 to 0.19 min, respectively. M. morganii had higher D-values than P. phosphoreum at the same pressure, indicating it was more resistant to HHP treatment. HFB had a higher D-value in tuna meat slurry compared with that in phosphate buffer, indicating that the HFB were more resistant to pressure in tuna meat slurry. The Zp values (pressure range that results in a 10-fold change in D-value) of M. morganii and P. phosphoreum were 162 and 140 MPa in phosphate buffer and 153 and 105 MPa in tuna meat slurry, respectively. Damage to the cell wall and cell membrane by HHP treatments can be observed by scanning electron microscopy. To our knowledge, this is the first report to demonstrate that HHP can be applied to inactivate the HFB M. morganii and P. phosphoreum by inducing morphological changes in the cells.

Inhibitory effect of a combination with novel jumbo bacteriophages ΦMV-1 and ΦMV-4 on Morganella morganii subsp. morganii growth and histamine accumulation.

Histamine (scombroid) poisoning is a foodborne illness caused by ingestion of histamine-contaminated seafood; therefore, inhibition of the growth of histamine-producing bacteria is key for it prevention. Infection of pathogenic bacteria by bacteriophages (phages) is being developed to prevent multiple foodborne illnesses. Here, we describe the inhibitory effect of a phage mixture on growth and histamine accumulation of Morganella morganii subsp. morganii, the primary causative agent of histamine poisoning in fish meat. We isolated novel two phages, ΦMV-1 and ΦMV-4, which infected M. morganii subsp. morganii strains tested in this study. ΦMV-1 and ΦMV-4 belong to family Myoviridae. Pulsed-field gel electrophoresis revealed that these phages are jumbo bacteriophages with large genomes. The latent period, rise period and burst size of ΦMV-1 were 30 min, 60 min, and 224 PFU per infected cell, respectively, and those of ΦMV-4 were 60 min, 50 min, and 62 PFU per infected cell, respectively. A mixture of ΦMV-1 and ΦMV-4 effectively prevented regrowth of M. morganii subsp. morganii after phage treatment, suggesting that the phage mixture treatment is more effective for inhibition of growth and histamine accumulation by M. morganii subsp. morganii than single phage treatment. Treatment with phage mixture inhibited growth and histamine accumulation by M. morganii subsp. morganii in canned and fresh tuna. The phage mixture might be an effective way to prevent growth of the histamine producer and accumulation of histamine in seafood.

A Rare Opportunist, Morganella morganii, Decreases Severity of Polymicrobial Catheter-Associated Urinary Tract Infection.

Catheter-associated urinary tract infections (CAUTIs) are common hospital-acquired infections and frequently polymicrobial, which complicates effective treatment. However, few studies experimentally address the consequences of polymicrobial interactions within the urinary tract, and the clinical significance of polymicrobial bacteriuria is not fully understood. Proteus mirabilis is one of the most common causes of monomicrobial and polymicrobial CAUTI and frequently cocolonizes with Enterococcus faecalis, Escherichia coli, Providencia stuartii, and Morganella morganiiP. mirabilis infections are particularly challenging due to its potent urease enzyme, which facilitates formation of struvite crystals, catheter encrustation, blockage, and formation of urinary stones. We previously determined that interactions between P. mirabilis and other uropathogens can enhance P. mirabilis urease activity, resulting in greater disease severity during experimental polymicrobial infection. Our present work reveals that M. morganii acts on P. mirabilis in a contact-independent manner to decrease urease activity. Furthermore, M. morganii actively prevents urease enhancement by E. faecalis, P. stuartii, and E. coli Importantly, these interactions translate to modulation of disease severity during experimental CAUTI, predominantly through a urease-dependent mechanism. Thus, products secreted by multiple bacterial species in the milieu of the catheterized urinary tract can directly impact prognosis.

Isolation, identification and characterization of Morganella morganii from Naja naja atra in Beijing, China.

Morganella morganii is an important opportunistic human pathogen and belongs to the family of Enterobacteriaceae. Although it is widely distribution, it only be considered a rare cause of human infections. We report the isolate of M. morganii from Naja naja atra following infections of heart, lung and liver. Seven strains were confirmed using 16S rDNA amplified and sequences. Antimicrobial susceptibility testing of M. morganii isolates demonstrated ubiquitous resistance to ampicillin, amoxicillin/clavulanic acid, cefazolin, cephalothin, sulfamethoxazole/trimethoprim, sulfamethoxazole et al. However, M. morganii ubiquitous susceptible to piperacillin, ampicillin/sulbactam, piperacillin/tazobactam, cefixime et al. Further investigate display gyr B and Sul2 genes presence in all M. morganii isolates. AAC(3)-II was found in E2, E3 and E6 M. morganii. gyrA and qnrB expression in M3 and M6 M. morganii. This is the first description in M. morganii carrying AAC(3)-II, gyrB, gyrA, qnrB, and Sul2 genes from Naja naja atra, which suggests the increasing risk of pathogen transmission between humans and wildlife.

Development of a real-time PCR method coupled with a selective pre-enrichment step for quantification of Morganella morganii and Morganella psychrotolerans in fish products.

Histamine fish poisoning is common and due to toxic concentrations of histamine often produced by Gram-negative bacteria in fin-fish products with a high content of the free amino acid histidine. The genus Morganella includes two species previously reported to cause incidents of histamine fish poisoning. Morganella morganii and Morganella psychrotolerans are both strong producer of histamine. However, little is known about the occurrence and critical stages for fish contamination with these bacteria. To elucidate contamination routes of Morganella, specific real-time quantitative PCR (RTi qPCR) methods for quantification of M. morganii and M. psychrotolerans have been developed. Selective primers amplified a 110 bp region of the vasD gene for M. psychrotolerans and a 171 bp region of the galactokinase gene for M. morganii. These primer-sets showed high specificity as demonstrated by using purified DNA from 23 other histamine producing bacteria and 26 isolates with no or limited histamine production. The efficiency of the qPCR reactions on artificially contaminated fish samples were 100.8% and 96.3% respectively. The limit of quantification (LOQ) without enrichment was 4 log CFU/g. A quantitative enrichment step with a selective medium was included and improved the sensitivity of the methods to a LOQ of below 50 CFU/g in seafood. RTi qPCR methods with or without enrichment were evaluated for enumeration of Morganella species in naturally contaminated fresh fish and lightly preserved seafood from Denmark. These new methods will contribute to a better understanding of the occurrence and histamine production by Morganella species in fish products, information that is essential to reduce the unacceptably high frequency of histamine fish poisoning.

Protein content analysis and antimicrobial activity of the crude venom of Montivipera bornmuelleri; a viper from Lebanon.

Viperidae snakes venoms represent a source of efficient bioactive components that have already led to the development of several new drugs. In this work, we analyzed the protein content of the Montivipera bornmuelleri crude venom using LC-ESI-MS, sephadex G-75 gel filtration and SDS-PAGE and demonstrated the presence of proteins with molecular masses corresponding to metalloprotease III, serine-protease and PLA2 in three fractions collected after gel filtration. Equally, we examined the antimicrobial effect of the venom that showed an important potency, as bactericidal agent, based on MBC and MIC values obtained, against Staphylococcus aureus and Morganella morganii bacteria. However, no activity was registered against Enterococcus faecalis, being the most resistant bacteria, neither against Aspergillus flavus and Penicillium digitatum fungal. Furthermore, on eleven other bacterial strains and the Candida albicans fungus, the venom has shown an intermediate efficacy by slightly reducing the growth. Our data concerning the Montivipera bornmuelleri venom give evidence of a rich and complex content aiding the exploration of new bioactive molecules for biopharmaceuticals purposes.

Heat resistance of histamine-producing bacteria in irradiated tuna loins.

Consumption of foods high in biogenic amines leads to an illness known as histamine, or scombrotoxin, poisoning. The illness is commonly associated with consumption of fish with high levels of histamine ( $ 500 ppm). The objective of this study was to determine and compare the heat resistance of five histamine-producing bacteria in irradiated albacore tuna loins. Heat-resistance parameters (D- and z-values) were determined for Morganella morganii, Raoultella planticola, Hafnia alvei, and Enterobacter aerogenes. D- or z-values were not determined for Photobacterium damselae, which was the most heat-sensitive organism in this study. P. damselae declined > 5.9 log CFU/g after a heat treatment of 50°C for 10 min, 54°C for 3 min, and 56°C for 0.5 min. M. morganii was the most heat-resistant histamine-producing bacteria in albacore tuna loins, followed by E. aerogenes, H. alvei, and R. planticola. M. morganii and E. aerogenes had the highest D(50°C), 49.7 ± 17.57 and 51.8 ± 17.38 min, respectively. In addition, M. morganii had the highest D-values for all other temperatures (54, 56, and 58°C) tested. D- and zvalues were also determined for M. morganii in skipjack tuna. While no significant (P > 0.05) difference was observed between D(54°C) and D(56°C) of M. morganii in either albacore or skipjack tuna, the D(58°C) (0.4 ± 0.17 min) was significantly lower (P < 0.05) in skipjack than in albacore (0.9 ± 0.24 min). The z-values for all organisms tested were in the range of 3.2 to 3.8°C. This study suggests that heat treatment designed to control M. morganii in tuna loins is sufficient for controlling histamine-producing bacteria in canned-tuna processing environments.

Growth, inactivation and histamine formation of Morganella psychrotolerans and Morganella morganii - development and evaluation of predictive models.

Mathematical models for growth, heat inactivation and histamine formation by Morganella psychrotolerans and Morganella morganii were studied to evaluate the importance of these bacteria in seafood. Curves for growth and histamine formation by M. psychrotolerans in broth and seafood were generated at constant and changing storage temperatures (n=12). Observed and predicted times to formation of 100, 500 and 2000 ppm histamine were used for evaluation of an existing M. psychrotolerans histamine formation model [Emborg, J., Dalgaard, P., 2008-this issue-this issue. Modelling and predicting the growth and histamine formation by Morganella psychrotolerans. International Journal of Food Microbiology. doi:10.1016/j.ijfoodmicro.2008.08.016] Growth rates for M. psychrotolerans and M. morganii were determined at different constant temperatures from 0 degrees C to 42.5 degrees C whereas heat inactivation was studied between 37.5 degrees C and 60 degrees C. A M. morganii growth and histamine formation model was developed by combining these new data (growth rate model) and data from the existing literature (maximum population density and yield factor for histamine formation). The developed M. morganii model was evaluated by comparison of predicted growth and histamine formation with data from the existing literature. Observed and predicted growth rates for M. psychrotolerans, at constant temperatures, were similar with bias- and accuracy factor values of 1.15 and 1.45, respectively (n=11). On average times to formation of critical concentrations of histamine by M. psychrotolerans were acceptably predicted but the model was not highly accurate. Nevertheless, predictions seemed useful to support decisions concerning safe shelf-life in relation to formulation, storage and distribution of chilled seafood. Parameters for the effect of temperature on growth and inactivation of M. psychrotolerans and M. morganii differed markedly with Tmin of -8.3 to -5.9 degrees C vs. 0.3 to 2.8 degrees C, Topt of 26.0 to 27.0 degrees C vs. 35.9 to 37.2 degrees C and Tmax 32.0 to 33.3 degrees C vs. 44.0 to 47.4 degrees C, D(50 degrees C) of 5.3 min vs. 13.1 min and z-values of 6.8 degrees C and 7.2 degrees C. At temperatures above approximately 15 degrees C M. morganii grew faster than M. psychrotolerans. Bias- and accuracy factor-values of 1.41 and 2.44 (n=93) showed the predicted growth of M. morganii to be faster than previously observed in fresh fish and broth. In agreement with this, predicted times to formation of critical histamine concentrations by M. morganii were on average shorter than observed in fresh fish. A combined model was suggested to predict histamine formation by both psychrotolerant and mesophilic Morganella during storage of fresh fish between 0 degrees C and 37 degrees C.

Biochemical and molecular characterization of three new variants of AmpC beta-lactamases from Morganella morganii.

Morganella morganii produces an inducible, chromosomally encoded AmpC beta-lactamase. We describe in this study three new variants of AmpC within this species with apparent pIs of 6.6 (M19 from M. morganii strain PP19), 7.4 (M29 from M. morganii strain PP29), and 7.8 (M37 from M. morganii strain PP37). After gene sequencing, deduced amino acid sequences displayed one to six substitutions when compared to the available Morganella AmpC sequences. An AmpR-encoding gene was also found upstream of ampC, including the LysR regulators' helix-turn-helix DNA-binding domain and the putative T-N11-A-protected region in the ampR-ampC intercistronic sequence. All three AmpC variants were purified from in vitro-generated derepressed mutants and showed overall similar kinetic parameters. None of the observed amino acid changes, occurring at the surface of the protein, appear to have a major influence in their catalytic properties. Morganella AmpCs exhibit the highest catalytic efficiencies (k(cat)/K(m)) on classical penicillins, cefoxitin, narrow-spectrum cephalosporins, and cefotaxime. Cefotaxime was more effectively hydrolyzed than other oxyimino-cephalosporins, whereas cefepime was 3 log-fold less efficiently hydrolyzed than other cephalosporins such as cephalothin. Several differences with other AmpC beta-lactamases were found. Ampicillin was more efficiently hydrolyzed than benzylpenicillin. High k(cat)/K(m) values were observed for oxacillin and piperacillin, which are usually poor substrates for AmpC. A fairly efficient hydrolysis of imipenem was detected as well. Aztreonam, carbenicillin, and tazobactam were effective transient inactivators of these variants.

Significant histamine formation in tuna (Thunnus albacares) at 2 degrees C--effect of vacuum- and modified atmosphere-packaging on psychrotolerant bacteria.

Occurrence and importance of psychrotolerant histamine producing bacteria in chilled fresh tuna were demonstrated in the present study. The objective was to evaluate microbial formation of histamine and biogenic amines in chilled fresh tuna from the Indian Ocean and stored either vacuum-packed (VP) or modified atmosphere-packed (MAP). Firstly, biogenic amines and the dominating microbiota were determined in VP tuna involved in an outbreak of histamine fish poisoning in Denmark. Secondly, the microbiota of fresh MAP tuna was evaluated at the time of processing in Sri Lanka and chemical, microbial and sensory changes were evaluated during storage at 1-3 degrees C. To explain the results obtained with naturally contaminated tuna the effect of VP and MAP on biogenic amine formation by psychrotolerant bacteria was evaluated in challenge tests at 2 degrees C and 10 degrees C. The VP tuna that caused histamine fish poisoning had a histamine concentration of >7000 mg/kg and this high concentration was most likely produced by psychrotolerant Morganella morganii-like bacteria or by Photobacterium phosphoreum. Similar psychrotolerant M. morganii-like bacteria dominated the spoilage microbiota of fresh MAP tuna with 60% CO2/40% N2 and formed >5000 mg/kg of histamine after 24 days at 1.7 degrees C. These psychrotolerant bacteria were biochemically similar to M. morganii subsp. morganii and their 16S rDNA (1495 bp) showed >98% sequence similarity to the type strain of this species. Toxic concentrations of histamine were produced at 2.1 degrees C in inoculated VP tuna by both the psychrotolerant M. morganii-like bacteria (7400+/-1050 mg/kg) and P. phosphoreum (4250+/-2050 mg/kg). Interestingly, MAP with 40% CO2/60% O2, in challenge tests, had a strong inhibitory effect on growth and histamine formation by both the psychrotolerant M. morganii-like bacteria and P. phosphoreum. In agreement with this, no formation of histamine was found in naturally contaminated fresh MAP tuna with 40% CO2/60% O2 during 28 days of storage at 1.0 degrees C. To reduce current problems with histamine fish poisoning due to VP tuna it is suggested, for lean tuna loins, to replace vacuum packaging with MAP containing approximately 40% CO2 and approximately 60% O2.

B1 cells contribute to serum IgM, but not to intestinal IgA, production in gnotobiotic Ig allotype chimeric mice.

B1 cells are a significant source of natural serum IgM, thereby serving as a first line of defense against systemic bacterial and viral infections. They can migrate to the intestinal lamina propria and differentiate into IgA-producing plasma cells and thus might play a similar role in mucosal immunity. To investigate the contribution of B1 cells to the intestinal IgA response induced by the commensal flora in immunocompetent animals, we generated gnotobiotic and conventionally reared Ig allotype chimeric mice. In this system B1- and B2-derived Abs can be distinguished based on different allotypes. FACS analysis of peritoneal cavity cells and analysis of B1- and B2-derived serum IgM indicated stable B1/B2 chimerism and the establishment of a functional B1 population. Monoassociation with either Morganella morganii, Bacteroides distasonis, or segmented filamentous bacteria induced germinal center reactions in Peyer's patches and led to the production of intestinal IgA, partially reactive with bacterial Ag. A considerable amount of serum IgM was B1 cell derived in both monoassociated and conventionally reared mice. However, most of the total as well as bacteria-specific intestinal IgA was produced by B2 cells. These data suggest that intestinal IgA production induced by commensal bacteria is mainly performed by B2, not B1, cells.

Morganella morganii causing solitary liver abscess complicated by pyopericardium and left pleural effusion in a nondiabetic patient.

Morganella morganii is a rare cause of solitary liver abscess in Taiwan. The complication of pyopericardium and pleural effusion in nondiabetic patient with solitary liver abscess are also rare. We present a case of a 48-year-old nondiabetic woman who experienced with epigastric discomfort 1 month prior to admission. Chills and fever developed 2 weeks before admission. Physical examination on admission revealed engorgement of the jugular vein over the right neck, precordial friction rubs, and tenderness over the right upper quadrant of abdomen. Chest film showed mild cardiomegaly and left pleural effusion. Computed tomography of the abdomen showed liver abscess, left hepatic lobe, pyopericardium, and left pleural effusion. M. morganii was isolated from 2 sets of blood cultures, one set of hepatic pus culture, and one set of pericardial pus culture. After pigtail drainage of liver abscess, pyopericardium for 12 days, and ceftriaxone intravenous administration for 19 days, the patient was discharged in stable condition.

New synthetic catecholate-type siderophores with triamine backbone.

New analogues of triscatecholate siderophores based on linear or tripodal triamines with or without spacer groups or lipophilic and hydrophilic substituents were synthesized. The catecholate moieties were prepared in OH-forms, as acetylated compounds or masked as 8-methoxycarbonyloxy-2,4-dioxo-1,3-benzoxazine derivatives. Some of the new compounds were active as siderophores tested by growth promotion assays using various gram-negative bacteria and mycobacteria under iron limitation and by CAS-assay. Structure-activity-correlations have been studied.

Human V gamma 2V delta 2 T cells produce IFN-gamma and TNF-alpha with an on/off/on cycling pattern in response to live bacterial products.

Whereas cytokine production in alphabeta T cells is rapidly regulated by exposure to peptide Ag, the mechanisms regulating cytokine production by gammadelta T cells are unknown. In this study, we demonstrate that human Vgamma2Vdelta2 T cells produce IFN-gamma and TNF-alpha as early as 2 h after Ag exposure, and that they produce these cytokines in a dose- and time- dependent manner in response to stimulation with a live bacterial product, iso-butylamine (IBA), but not to dead bacteria or LPS. gammadelta T cells began, ceased, and then resumed IFN-gamma and TNF-alpha generation in an on/off/on cycling pattern, both in vitro and in vivo, depending on the presence or absence of IBA. IFN-gamma and TNF-alpha, whose optimum production was dependent on IBA-stimulated gammadelta T cells, were critical for monocyte-mediated killing of Escherichia coli. By limiting cytokine production to periods of direct contact with live bacteria, gammadelta T cells focus their resources at the site of infection, while limiting systemic immunopathology. Thus, human gammadelta T cells may mediate innate resistance to extracellular bacteria via tightly regulated cytokine production without necessarily expanding in number.

Immunohistopathologic demonstration of pleuropneumonia associated with Morganella morganii in a piglet.

Serofibrinous pleuropneumonia in a piglet was examined microbiologically and immunohistopathologically. Large numbers of Morganella morganii were isolated from the pneumonic lesion, but no other pathogens were identified. A large amount of M. morganii antigen was demonstrated, and its distribution was closely associated with the histologic lesion. This finding suggests that pleuropneumonia in piglets might be caused by M. morganii.

Growth and histamine formation of Morganella morganii in determining the safety and quality of inoculated and uninoculated bluefish (Pomatomus saltatrix).

The objective of this study was to determine the effect of normal microflora and Morganella morganii on histamine formation and olfactory acceptability in raw bluefish under controlled storage conditions. Fillets inoculated with and without M. morganii were stored at 5, 10, and 15 degrees C for 7 days. Microbial isolates from surface swabs were identified and screened for histidine decarboxylase activity. Olfactory acceptance was performed by an informal sensory panel. Histamine levels were quantified using high-performance liquid chromatography and fluorescence detection. While olfactory acceptance decreased, histamine concentration and bacterial counts increased. Storage temperature had a significant effect on histamine levels, bacterial counts, and olfactory acceptance of the bluefish. Inoculation with M. morganii had a positive significant effect on histamine formation for bluefish held at 10 and 15 degrees C (P < 0.0001). The results of the study will serve in supporting U.S. Food and Drug Administration (FDA) regulations regarding guidance and hazard levels of histamine in fresh bluefish.

Histamine and biogenic amine production by Morganella morganii isolated from temperature-abused albacore.

Histamine-producing bacteria were isolated from albacore stored at 0, 25, 30, and 37 degrees C. They were screened using Niven's differential medium, and their histamine production was confirmed by high-pressure liquid chromatography analysis. The optimum temperature for growth of histamine-producing bacteria was 25 degrees C. The bacterium producing the highest level of histamine was isolated from fish abused at 25 degrees C. It was identified as Morganella morganii by morphological, cultural, biochemical, and antimicrobial characteristics and by the Vitek microbial identification system. The M. morganii isolate was inoculated into tuna fish infusion broth medium, and the effect of temperature was determined for microbial growth and formation of histamine and other biogenic amines. The isolate produced the highest level of histamine, 5,253 ppm, at 25 degrees C in the stationary phase. At 15 degrees C, histamine production was reduced to 2,769 ppm. Neither microbial growth nor histamine formation was detected at 4 degrees C. To determine whether the isolate can also produce other biogenic amines that can potentiate histamine toxicity, production of cadaverine, putrescine, serotonin, tryptamine, tyramine, phenylethylamine, spermidine, and spermine by the isolate was also monitored. Cadaverine, putrescine, and phenylethylamine were detected with microbial growth in the tuna fish infusion broth medium. The optimum temperature for cadaverine, putrescine, and phenylethylamine formation was found to be 25 degrees C, as it was for histamine.